Kshitij Srivastava1, Anne-Sophie Fratzscher1, Bo Lan1, Willy Albert Flegel2. 1. Laboratory Services Section, Department of Transfusion Medicine, NIH Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA. 2. Laboratory Services Section, Department of Transfusion Medicine, NIH Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA. waf@nih.gov.
Abstract
BACKGROUND: Clinically effective and safe genotyping relies on correct reference sequences, often represented by haplotypes. The 1000 Genomes Project recorded individual genotypes across 26 different populations and, using computerized genotype phasing, reported haplotype data. In contrast, we identified long reference sequences by analyzing the homozygous genomic regions in this online database, a concept that has rarely been reported since next generation sequencing data became available. STUDY DESIGN AND METHODS: Phased genotype data for a 80.6 kb region of chromosome 1 was downloaded for all 2,504 unrelated individuals of the 1000 Genome Project Phase 3 cohort. The data was centered on the ACKR1 gene and bordered by the CADM3 and FCER1A genes. Individuals with heterozygosity at a single site or with complete homozygosity allowed unambiguous assignment of an ACKR1 haplotype. A computer algorithm was developed for extracting these haplotypes from the 1000 Genome Project in an automated fashion. A manual analysis validated the data extracted by the algorithm. RESULTS: We confirmed 902 ACKR1 haplotypes of varying lengths, the longest at 80,584 nucleotides and shortest at 1,901 nucleotides. The combined length of haplotype sequences comprised 19,895,388 nucleotides with a median of 16,014 nucleotides. Based on our approach, all haplotypes can be considered experimentally confirmed and not affected by the known errors of computerized genotype phasing. CONCLUSIONS: Tracts of homozygosity can provide definitive reference sequences for any gene. They are particularly useful when observed in unrelated individuals of large scale sequence databases. As a proof of principle, we explored the 1000 Genomes Project database for ACKR1 gene data and mined long haplotypes. These haplotypes are useful for high throughput analysis with next generation sequencing. Our approach is scalable, using automated bioinformatics tools, and can be applied to any gene.
BACKGROUND: Clinically effective and safe genotyping relies on correct reference sequences, often represented by haplotypes. The 1000 Genomes Project recorded individual genotypes across 26 different populations and, using computerized genotype phasing, reported haplotype data. In contrast, we identified long reference sequences by analyzing the homozygous genomic regions in this online database, a concept that has rarely been reported since next generation sequencing data became available. STUDY DESIGN AND METHODS: Phased genotype data for a 80.6 kb region of chromosome 1 was downloaded for all 2,504 unrelated individuals of the 1000 Genome Project Phase 3 cohort. The data was centered on the ACKR1 gene and bordered by the CADM3 and FCER1A genes. Individuals with heterozygosity at a single site or with complete homozygosity allowed unambiguous assignment of an ACKR1 haplotype. A computer algorithm was developed for extracting these haplotypes from the 1000 Genome Project in an automated fashion. A manual analysis validated the data extracted by the algorithm. RESULTS: We confirmed 902 ACKR1 haplotypes of varying lengths, the longest at 80,584 nucleotides and shortest at 1,901 nucleotides. The combined length of haplotype sequences comprised 19,895,388 nucleotides with a median of 16,014 nucleotides. Based on our approach, all haplotypes can be considered experimentally confirmed and not affected by the known errors of computerized genotype phasing. CONCLUSIONS: Tracts of homozygosity can provide definitive reference sequences for any gene. They are particularly useful when observed in unrelated individuals of large scale sequence databases. As a proof of principle, we explored the 1000 Genomes Project database for ACKR1 gene data and mined long haplotypes. These haplotypes are useful for high throughput analysis with next generation sequencing. Our approach is scalable, using automated bioinformatics tools, and can be applied to any gene.
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Authors: Grace X Y Zheng; Billy T Lau; Michael Schnall-Levin; Mirna Jarosz; John M Bell; Christopher M Hindson; Sofia Kyriazopoulou-Panagiotopoulou; Donald A Masquelier; Landon Merrill; Jessica M Terry; Patrice A Mudivarti; Paul W Wyatt; Rajiv Bharadwaj; Anthony J Makarewicz; Yuan Li; Phillip Belgrader; Andrew D Price; Adam J Lowe; Patrick Marks; Gerard M Vurens; Paul Hardenbol; Luz Montesclaros; Melissa Luo; Lawrence Greenfield; Alexander Wong; David E Birch; Steven W Short; Keith P Bjornson; Pranav Patel; Erik S Hopmans; Christina Wood; Sukhvinder Kaur; Glenn K Lockwood; David Stafford; Joshua P Delaney; Indira Wu; Heather S Ordonez; Susan M Grimes; Stephanie Greer; Josephine Y Lee; Kamila Belhocine; Kristina M Giorda; William H Heaton; Geoffrey P McDermott; Zachary W Bent; Francesca Meschi; Nikola O Kondov; Ryan Wilson; Jorge A Bernate; Shawn Gauby; Alex Kindwall; Clara Bermejo; Adrian N Fehr; Adrian Chan; Serge Saxonov; Kevin D Ness; Benjamin J Hindson; Hanlee P Ji Journal: Nat Biotechnol Date: 2016-02-01 Impact factor: 54.908